title: ENTPD3 Marks Mature Stem Cell Derived Beta Cells Formed by Self-Aggregation in Vitro

city: Aurora

first: Kent

middle: Augustus

last: Riemondy

state: Colorado

department: Biochemistry and Molecular Genetics

institute: University of Colorado School of Medicine

summary: Stem cell derived beta-like cells (sBC) carry the promise of providing an abundant source of insulin-producing cells for use in cell replacement therapy for patients with diabetes, potentially allowing widespread implementation of a practical cure. To achieve their clinical promise, sBC need to function comparably to mature adult beta cells, but as yet they display varying degrees of maturity. Indeed, detailed knowledge of the events resulting in human beta cell maturation remains obscure. Here we show that sBC spontaneously self-enrich into discreet islet-like cap structures within in vitro cultures, independent of exogenous maturation conditions. Multiple complementary assays demonstrate that this process is accompanied by functional maturation of the selfenriched sBC (seBC); however, the seBC still contain distinct subpopulations displaying different maturation levels. Interestingly, the surface protein ENTPD3 (also known as nucleoside triphosphate diphosphohydrolase-3 (NDPTase3)) is a specific marker of the most mature seBC population and can be used for mature seBC identification and sorting. Our results illuminate critical aspects of in vitro sBC maturation and provide important insights towards developing functionally mature sBC for diabetes cell replacement therapy.

overall_design: Single cell RNA-Seq and bulk RNA-seq of human iPSC induced beta cells

first: Fiona

middle: M

last: Docherty

first: Jay

last: Hesselberth

first: Holger

last: Russ


title: A Systems Approach to Interrogate Gene Expression Patterns in African American Men Presenting with Clinically Localized Prostate Cancer

status: Public on Oct 12 2021

city: Charleston

first: Gary

last: Hardiman

state: SC

address: 135 Cannon Street

department: Medicine

country: USA

institute: Medical University of South Carolina

type: Expression profiling by high throughput sequencing

summary: In this study, we used a precision medicine approach to examine molecular profiles and cellular stress responses in an important segment of AA and EA men: men undergoing prostate biopsy.  The primary goal in the present study was to demonstrate the ability to assess the prostate transcriptome with a single biopsy core while further examining molecular differences in gene expression patterns relevant to prostate cancer disparities between AA and EA men in a new patient cohort presenting with early stage prostate cancer. To further this objective, we undertook a series of genome wide expression profiling experiments using high throughput RNA sequencing (RNA-Seq). RNA was purified from core biopsy prostate tissue specimens obtained from deidentified subjects enrolled in the study. Transcriptional profiles of each of the deidentified patient’s tissue samples were generated and systems level analyses were performed. The results of these analyses were compared across racial groups.

overall_design: RNA-seq of prostate biopsy from AA and EA men with low or high vitamin D serum level.

first: Stephen

middle: J

last: Savage

first: E

middle: S

last: Hazard

first: Willian

middle: A

last: da Silveira

first: Robert

middle: C

last: Wilson

first: Susan

last: Caulder

first: Linda

last: Ambrose

first: Lewis

last: Frey

first: Bethany

last: Wolf

first: Sebastiano

last: Gattoni-Celli

first: Chanita

last: Hughes Halbert


title: Immunomodulatory cytokines control the expression of a leukemic state in a de novo model of MYC-induced human AML

city: Vancouver

first: Elizabeth

last: Bulaeva

state: BC

address: 675 West 10th Avenue

department: Terry Fox Laboratory

country: Canada

institute: British Columbia Cancer Agency

summary: Human acute myeloid leukemias (AMLs) comprise a genetically and biologically diverse group of clonal hematologic malignancies. Nevertheless, they all share the defining features of deregulated growth and a blockade of normal terminal differentiation events. Hematopoietic stem cells appear to be the origin of clones from which human AML may often arise, but the intrinsic and extrinsic processes that establish self-sustaining, overtly leukemic human cell populations remain poorly understood, exacerbated by historical difficulties in developing de novo models of AML in human cells and drawbacks inherent in analyzing leukemic cells obtained from patients or model organisms. The finding that MYC expression is commonly increased in AML cells carrying different upstream driver mutations led us to examine the effects of forcing a modestly enhanced level of expression of this gene in freshly isolated human CD34+ hematopoietic cells. We now show that both early and late types of cells with granulopoietic potential when transduced with a MYC-encoding lentiviral vector produce a rapidly fatal, phenotypically and transcriptionally indistinguishable, serially transplantable AML at high efficiency in immunodeficient mice, but only in recipients that are producing human interleukin-3 (IL3) and/or granulocyte-macrophage colony-stimulating factor (GM-CSF). In the absence of these human growth factors (GFs), transplants of MYC-transduced CD34+ cells regenerate a normal spectrum of human lymphoid and myeloid progeny. However, upon secondary transfer to human GF-producing mice, they produce a phenotypically similar, rapidly fatal AML. The unique features of this model that include the reproducible creation of a latent AML program in primary human cells that is rapidly activated by inflammatory growth factors portend its utility as a powerful new platform for elucidating critical shared molecular mechanisms responsible for creating aggressive human myeloid leukemias.

overall_design:
bulk RNA-seq: 6 AML generated in NRG-3GS mice by injecting CD34+CD38- or GMP cord blood cells lentivirally transduced with a vector carrying the cDNA for MYC. At Euthanasia, bone marrow or spleen cells were flow sorted for humanCD33 and GFP prior to RNA extraction.

single cell RNA-seq: 4 AML generated in NRG-3GS mice by injecting CD34+CD38- or GMP cord blood cells lentivirally transduced with a vector carrying the cDNA for MYC. At Euthanasia, spleen cells were flow sorted for human CD33 and GFP prior to RNA extraction.

first: Davide

last: Pellacani

first: Colin

last: Hammond

first: Michelle

last: Moksa

first: Annaick

last: Carles

first: Martin

last: Hirst

first: Connie

last: Eaves


title: Analysis of Scavenger-Receptor Type H deficient liver sinusoidal endothelial cells

status: Public on Oct 11 2021

city: Mannheim

first: Carsten

last: Sticht

address: Theodor-Kutzer-Ufer

department: ZMF

country: Germany

institute: University Heidelberg

summary:
Two of the most highly abundant transcripts in liver sinusoidal endothelial cells are Stab1 and Stab2, we investigated downstream effects on liver sinusoidal endothelial cells by disrupting their expression.
We used microarrays to detail the global programme of gene expression in liver sinusoidal endothelial cells deficient for Scavenger-Receptor type H compared to Wildtype.

overall_design: Mice aged 3-4 months (WT, Stab1KO, Stab2KO, Stab1/2DKO) were sacrificed, the liver was perfused and liver sinusoidal endothelial cells were isolated using CD146 magnetic beads. RNA was isolated immediately afterwards and prepared for microarray analysis.

first: Philipp-Sebastian

last: Koch


title: The role of microRNA-204 in pressure overload-induced cardiac hypertrophy

city: Iowa City

first: Ajit

last: Vikram

state: IA

address: 285 Newton Road

department: Internal Medicine

institute: University of Iowa

summary: microRNAs regulate cardiac hypertrophy development, which predicts the risk of heart failure. Here we investigate the role of microRNA-204-5p (miR-204) in developing cardiac hypertrophy and cardiac dysfunction following transaortic constriction. To determine the role of miR-204, we determined the transcriptomic profile of hearts following transaortic constriction.

overall_design: The wildtype or miR-204-/- mice were subjected to transaortic constriction surgery, and heart samples were collected five weeks after surgery.


title: Role of BTK and PI3K-P110d in JMML

city: INDIANAPOLIS

first: Xiaoyu

last: Lu

state: IN

address: 410W 10TH ST

department: school of medicine

institute: INDIANA UNIVERSITY

summary: Individual drug treatment in a mouse model of JMML equally ameliorates monocytosis and splenomegaly but combined treatment uniquely ameliorates thrombocytopenia.

overall_design: To elucidate the gene expression changes impacted on BTK and  PI3K-P110δ treatment in Shp2 E76K mice

first: Reuben

last: Kapur


title: Circadian regulation of SARS-CoV-2 infection in lung epithelial cells

city: Oxford

first: Xiaodong

last: Zhuang

state: Oxfordshire

address: Old Road Campus, Roosevelt Drive

country: United Kingdom

institute: University of Oxford

summary: Calu-3 cells with stable shBmal1 silencing or SR9009 treatment were performed RNA-seq analysis. The sequence reads were mapped to the human genome.

overall_design: mRNA profiles of control, Bmal1 silenced and SR9009 treated Calu-3 cells

first: Jane

last: McKeating


title: Differential effects of two catalytic mutations on full-length PRDM9 and its isolated PR/SET domain reveal a case of pseudo-modularity

city: Bar Harbor

first: Natalie

middle: R

last: Powers

state: ME

address: 600 Main Street

department: Research

institute: The Jackson Laboratory

type: Genome binding/occupancy profiling by high throughput sequencing

summary: The locations of mammalian recombination hotspots are determined by PRDM9, a zinc finger histone methyltransferase that locally trimethylates histone H3 at residues K4 and K36. We previously reported two hypomorphic catalytic mutations, Prdm9-EP and Prdm9-EK, with different phenotypic effects. Prdm9-EP, but not Prdm9-EK, is compatible with female sub-fertility, while both mutations phenocopy the Prdm9-null condition in males. Here we directly compare and contrast the enzymatic effects of the two mutations in vitro and in vivo. We previously performed two biological H3K4me3 ChIP-seq replicates in spermatocytes isolated from Prdm9-EP homozygous males (GSE144144; SRX8588740 and SRX8588741), and re-processed previously reported H3K4me3 ChIP-seq data from spermatocytes isolated from wild-type B6 males (GSE52628; SRX381465 and SRX381466). We used those raw and processed files for this study (GSE144144). We also previously performed one biological H3K4me3 replicate in spermatocytes isolated from Prdm9-EP homozygous males (GSE112110; SRX4136625). We report an additional replicate here, and merged the two replicates for analysis; raw and processed files are reported here. We also performed ChIP-seq for H3K36me3 in both Prdm9-EP and Prdm9-EK homozygous spermatocytes. Raw and processed files are available here. For comparison, we re-mapped and re-analyzed H3K36me3 ChIP-seq data we previously reported from wild-type B6 spermatocytes (GSE76416; SRX1508234); processed files are available here.

overall_design: We performed ChIP-seq with antibodies against H3K4me3 and H3K36me3, using chromatin extracted from C57BL/6J-Prdm9-EP and C57BL/6J-Prdm9-EK homozygous mutant spermatocytes, and combined it with previously generated ChIP-seq data for H3K4me3 in C57BL/6J wild-type and C57BL/6J-Prdm9-EP spermatocytes, for direct comparison between the three genotypes.

first: Petko

last: Petkov


title: CDK8 is associated with RAP2.6 and SnRK2.6 and positively modulates abscisic acid signaling and drought response in Arabidopsis

city: West Lafayette

first: Jian-Kang

last: Zhu

address: 625 Agriculture Mall Dr.

department: Department of Horticulture and Landscape Architecture

institute: Purdue University

summary: CDK8 is a key subunit of Mediator complex, a large multiprotein complex that is a fundamental part of the conserved eukaryotic transcriptional machinery. However, the biological functions of CDK8 in plant abiotic stress responses remain largely unexplored. Here, we demonstrated CDK8 as a critical regulator in the abscisic acid (ABA) signaling and drought response pathways in Arabidopsis. Compared to wild-type, cdk8 mutants showed reduced sensitivity to ABA, impaired stomatal apertures and hypersensitivity to drought stress. Transcriptomic and chromatin immunoprecipitation analysis revealed that CDK8 positively regulates the transcription of several ABA-responsive genes, probably through promoting the recruitment of RNA polymerase II to their promoters. We discovered that both CDK8 and SnRK2.6 interact physically with an ERF/AP2 transcription factor RAP2.6, which can directly bind to the promoters of RD29A and COLD-REGULATED 15A (COR15A) with GCC or DRE elements, thereby promoting their expression. Importantly, we also showed that CDK8 is essential for the ABA-induced expression of RAP2.6 and RAP2.6-mediated upregulation of ABA-responsive genes, indicating that CDK8 could link the SnRK2.6-mediated ABA signaling to RNA polymerase II to promote immediate transcriptional response to ABA and drought signals. Overall, our data provide new insights into the roles of CDK8 in modulating ABA signaling and drought responses.

overall_design: RNA-Seq of 4 samples, each has 3 replicates. The samples are wild type ( WT ) with mock treatment, cdk8 with mock treatment, WT with 50 μM ABA treatment for 3 hours, and cdk8 with 50 μM ABA treatment for 3 hours.

first: Yingfang

first: Kai

last: Tang


title: Expression data from sorafenib-resistant HCC cells, xenograft model and the corresponding counterparts.

city: Suzhou

first: Yanmin

last: Xu

address: Suzhou industrial park, 199 renai road

department: Pathology Center and Department of Pathology

country: China

institute: Soochow University

summary: To better identify the key gene involved in sorafenib-resistant HCC cells and uncover potential targets for HCC therapy, the microarray analysis was used to screen the differentially expressed genes in sorafenib-resistant HCC cells, xenograft model and the corresponding counterparts.

overall_design: Parental or Sorafenib-resistant HCC cell lines and HCC xenografts were for RNA extraction and hybridization on Affymetrix microarrays.


title: Partitioning RNAs by length improves transcriptome reconstruction from short-read RNA-seq data

city: Munich

first: Francisca

last: Rojas Ringeling

address: Feodor-Lynen-Straße 25, 81377

department: Biochemistry

institute: Ludwig Maximilian University of Munich

summary: The accuracy of methods for assembling transcripts from short-read RNA sequencing data is limited by the lack of long-range information. Here we introduce Ladder-seq, an approach that separates transcripts according to their lengths prior to sequencing and uses the additional information to improve the quantification and assembly of transcripts. Using simulated data, we demonstrate that a kallisto algorithm extended to process Ladder-seq data quantifies transcripts of complex genes with substantially higher accuracy than conventional kallisto. For reference-based assembly, a modified StringTie2 algorithm reconstructs a single transcript with 30.8% higher precision than its conventional counterpart and is >30% more sensitive for complex genes. For de novo assembly, a modified Trinity algorithm correctly assembles 78% more transcripts than conventional Trinity, while improving precision by 78%. In experimental data, Ladder-seq reveals 40% more genes harboring isoform switches compared with conventional RNA-seq and unveils widespread changes in isoform usage upon m6A depletion by Mettl14 knock-out.

overall_design: RNA sequencing of  neural progenitor cells from Mettl14 wild type and conditional knock-out mice with four replicates per genotype. Prior to sequencing mRNA from each sample was separated by length into 7 distinct bands. Each band from each sample has a unique barcode.

last: Ringeling

first: Stefan

last: Canzar


title: 4DNESB7XYI9V - sci-Hi-C on mESCs differentiated to embryoid body

city: Boston

last: DCIC

state: MA

address: 10 Shattuck St

institute: 4D Nucleome - Data Coordination and Integration Center

summary:
Mammalian development is associated with extensive changes in gene expression, chromatin accessibility, and nuclear structure. Here, we follow such changes associated with mouse embryonic stem cell differentiation and X inactivation by integrating, for the first time, allele-specific data obtained by high-throughput single-cell RNA-seq, ATAC-seq, and Hi-C. In differentiated cells, contact decay profiles, which clearly distinguish the active and inactive X chromosomes, reveal loss of the inactive X-specific structure at mitosis followed by a rapid reappearance, suggesting a bookkeeping mechanism. In differentiating embryonic stem cells, changes in contact decay profiles are detected in parallel on both the X chromosomes and autosomes, suggesting profound simultaneous reorganization. The onset of the inactive X-specific structure in single cells is notably delayed relative to that of gene silencing, consistent with the idea that chromatin compaction is a late event of X inactivation. Novel computational approaches to effectively align single-cell gene expression, chromatin accessibility, and 3D chromosome structure reveal that long-range structural changes to chromosomes appear as discrete events, unlike progressive changes in gene expression and chromatin accessibility.

### Competing Interest Statement

The authors have declared no competing interest.

overall_design:
lab: Christine Disteche, UW
award: 1U54DK107979-01
accession: 4DNESB7XYI9V
submitted_by: Giancarlo Bonora
dataset_label: sci-Hi-C on mESCs differentiated to embryoid body
contributing_labs: William Noble, UW, Zhijun Duan, UW, Jay Ashok Shendure, UW
experiment_type: sci-Hi-C
url: https://data.4dnucleome.org/experiment-sets/4DNESB7XYI9V/
number_of_experiments: 23

experiment: 4DNEXOA4DNR8; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXCDN76F9; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXJ33QCZU; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXLCFXRUP; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEX1BY7H2C; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXU9FJ24U; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXTJOSWTW; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXMYV77X1; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXK2CP3J1; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXXNEYDTM; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXJNGKL32; replicate_number: Biorep 2, Techrep 1
experiment: 4DNEXJAHCEIN; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXLZWN9VH; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXV3C383J; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXB7EWKCM; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXTLVDZQP; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXEE1N4JI; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXPLD13JA; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXVG7LXZE; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXWHKWMSY; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXRMWXOKT; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXMN3ZJ4X; replicate_number: Biorep 1, Techrep 1
experiment: 4DNEXMNAHJPG; replicate_number: Biorep 2, Techrep 1

Series supplementary files:
***************
genome_assembly: GRCm38
description: A tarball containing processed sci-Hi-C data.
file name: 4DNFIZ8TEE2M.tar
file_format: tar
***************
file_type: readme
description: A text file describing the contents and processing steps for the sci-Hi-C processed data in 4DNFIZ8TEE2M.
file name: 4DNFI7QQWLOV.txt
file_format: txt
***************
file_type: directory tree
description: A text file with a directory tree for the sci-Hi-C processed data in 4DNFIZ8TEE2M.
file name: 4DNFICOPS6ER.txt
file_format: txt


title: A Polycomb domain found in committed cells impairs differentiation when introduced into PRC1 in pluripotent cells (EBs/NPCs RNA-seq datasets)

first: Robert

last: Kingston

address: 185 Cambridge Street, 7th Floor

department: Molecular Biology

institute: Massachusetts General Hospital

summary: The CBX family of proteins is central to proper mammalian development via key roles in Polycomb-mediated maintenance of repression. CBX proteins in differentiated lineages have chromatin compaction and phase separation activities that might contribute to maintaining repressed chromatin. The predominant CBX protein in pluripotent cells, CBX7, lacks the domain required for these activities. We inserted this functional domain into CBX7 in embryonic stem cells to test the hypothesis that it contributes a key epigenetic function. ESCs expressing this chimeric CBX7 were impaired in their ability to properly form embryoid bodies and neural progenitor cells and showed reduced activation of lineage-specific genes across differentiation. Neural progenitors exhibited a corresponding inappropriate maintenance of Polycomb binding at neural-specific loci over the course of differentiation. We propose that a switch in the ability to compact and phase separate is a central aspect of Polycomb group function during the transition from pluripotency to differentiated lineages.

overall_design: Embryonic stem cells expressing different versions of CBX7 (WT, CaPS, mCaPS, KO) were differentiated into embryoid bodies and neural progenitor cells. Gene expression profiles were determined for each cell type across differentiation by performing RNA-Seq on three replicates each.

first: Elizabeth

last: Jaensch

middle: E


title: Agilent zebrafish V3 array 4´44K microarray-Aflatoxin B1 treated with 6-48 hpf zebrafish embryos

city: Taipei

first: Ting-Shuan

last: Wu

address: 5F ,No 1, Sec 1, Jen-Ai Rd

country: Taiwan

institute: ntu

summary: To investigate the effects of Aflatoxin B1 on gene expressions in zebrafish embryos.

overall_design: Aflatoxin B1  mediated mRNA expression in zebrafish embryos which were exposed from 6 hpf to 48 hpf with doses of 0 and 150 ng/mL

first: Biing-Hui

last: Liu


title: Telomere-specific chromatin capture using a pyrrole–imidazole polyamide probe for the identification of proteins and non-coding  [PI-PRICh, DNA_pulldown2]

status: Public on Oct 10 2021

city: Mishima

first: Satoru

last: Ide

address: 1111 Yata

country: Japan

institute: National Institute of Genetics

summary:
Background: Knowing chromatin components at a DNA regulatory element at any given time is essential for understanding how the element works during cellular proliferation, differentiation and development. A region-specific chromatin purification is an invaluable approach to dissecting the comprehensive chromatin composition at a particular region. Several methods (e.g., PICh, enChIP, CAPTURE and CLASP) have been developed for isolating and analyzing chromatin components. However, all of them have some shortcomings in identifying non-coding RNA associated with DNA regulatory elements.
Results: We have developed a new approach for affinity purification of specific chromatin segments employing an N-methyl pyrrole (P)−N-methylimidazole (I) (PI) polyamide probe, which binds to a specific sequence in double-stranded DNA via Watson–Crick base pairing as a minor groove binder. This new technique is called proteomics and RNA-omics of isolated chromatin segments (PI-PRICh). Using PI-PRICh to isolate mouse and human telomeric components, we found enrichments of shelterin proteins, the well-known telomerase RNA component (TERC) and telomeric repeat-containing RNA (TERRA). When PI-PRICh was performed for alternative lengthening of telomere (ALT) cells with highly recombinogenic telomeres, in addition to the conventional telomeric chromatin, we obtained chromatin regions containing telomeric repeat insertions scattered in the genome and their associated RNAs.
Conclusion: PI-PRICh reproducibly identified both the protein and RNA components of telomeric chromatin when targeting telomere repeats. PI polyamide is a promising alternative to simultaneously isolate associated proteins and RNAs of sequence-specific chromatin regions under native conditions, allowing better understanding of chromatin organization and functions within the cell.

overall_design: DNA regions pulled down by TH59-DB pull-down and DNA regions of input in PI-PRICh for mouse erythrocytes leukemia cells

first: Asuka

last: Sasaki


title: Telomere-specific chromatin capture using a pyrrole–imidazole polyamide probe for the identification of proteins and non-coding [PI-PRICh, RNA_pulldown1]

overall_design: RNA profiles of input, masked TH59-DB pull-down, TH59-DB pull-down fraction of PI-PRICh for mouse erythrocytes leukemia cells


title: The lncRNA HERVH negatively regulates chromatin targeting and remodeling mediated by CHD7 [RNA-seq]

first: Fei

last: Ji

address: 185 Cambridge St

summary: Chromodomain helicase DNA-binding protein 7 (Chd7) encodes an ATP-dependent chromatin remodeler required for normal human development. De novo mutations in CHD7 are the major cause of CHARGE syndrome which features multiple developmental defects. Previous studies have shown that CHD7 targets to specific genomic loci and alters local transcription potentially by remodeling the chromatin structure in a cell stage and lineage specific manner. We examined whether nuclear RNAs might contribute to its targeting and function. We used PAR-CLIP to identify a lncRNA, HERVH, that is uniquely expressed in pluripotent stem cells and preferentially interacts with CHD7. Knockdown of HERVH using LNAs or knockout of an individual copy of HERVH by CRISPR-Cas9 both resulted in increased binding of CHD7 and increased levels of H3K27ac at a subset of enhancers. Depletion of HERVH led to the activation of multiple genes, several of which might potentially poise cells towards differentiation. CHD7 bound HERVH RNA with high affinity but low specificity and that this interaction decreased the ability of CHD7 to bind and remodel nucleosomes. We present a model in which HERVH acts as a decoy to modulate the dynamics of CHD7 binding to enhancers in pluripotent cells and to prevent the activation of differentiation specific genes.

overall_design: Examination of gene expression, CHD7 occpancy and histone modifications on chromatin in WT and HERVH depleted hESCs.

first: Fu-Kai

last: Hsieh

first: Manashree

last: Damle

first: Ruslan

middle: I

last: Sadreyev


title: RNA Binding Proteins As Regulators of Oxidative Stress Identified by a Targeted CRISPR-Cas9 Single Guide RNA Library

city: Cambridge

first: Louise

last: Matheson

address: Babraham Research Campus

department: Laboratory of Lymphocyte Signalling and Development

institute: The Babraham Institute

summary: The clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 genome editing system has been broadly adopted for high-throughput genetic screens. However, the application of genome-wide single guide RNA (sgRNA) libraries can be challenging. We generated a custom sgRNA library, an order of magnitude smaller than genome-wide alternatives, to facilitate the genetic screening of RNA binding proteins (RBPs). We demonstrated the utility of our reagent in a genetic screen for RBPs that conveyed cellular resistance or sensitivity to oxidative stress induced by paraquat. This identified that CSDE1 and STRAP, proteins that interact with each other, convey sensitivity to oxidative stress and that Pumilio homologues (PUM1 and PUM2) convey resistance. Targeting eIF4-E1 and -A1 protected cells from high-dose paraquat, whereas eIF4E2 targeted cells did less well. We also found that G3BP1 promoted sensitivity to a low dose of paraquat but protected cells at a higher dose. Our study highlights the use of genetic screens to identify roles of RBPs and identifies novel genes regulating sensitivity to oxidative stress.

overall_design: An sgRNA library targeting human RNA binding proteins was transduced into Jurkat T cells expressing Cas9. Guide representation was assessed by NGS following 14 or 21 days culture in 0, 50 or 200μM paraquat, with 1 replicate per condition. One replicate of cells at day 0 (without paraquat) was also included.

first: David

last: Turner

first: Martin


title: Telomere-specific chromatin capture using a pyrrole–imidazole polyamide probe for the identification of proteins and non-coding  [PI-PRICh, DNA_pulldown1]

overall_design: DNA regions pulled down by TH59-DB pull-down in PI-PRICh for HeLa1.3 and U2-OS cells


title: Telomere-specific chromatin capture using a pyrrole–imidazole polyamide probe for the identification of proteins and non-coding

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: The lncRNA HERVH negatively regulates chromatin targeting and remodeling mediated by CHD7 [PAR-CLIP]


title: The lncRNA HERVH negatively regulates chromatin targeting and remodeling mediated by CHD7 [CUT&RUN]


title: Telomere-specific chromatin capture using a pyrrole–imidazole polyamide probe for the identification of proteins and non-coding  [PI-PRICh, RNA_pulldown2]

overall_design: RNA profiles of input, masked TH59-DB pull-down, TH59-DB pull-down fraction of PI-PRICh for HeLa1.3 and U2-OS cells


title: The lncRNA HERVH negatively regulates chromatin targeting and remodeling mediated by CHD7 [ATAC-seq]


title: Single-cell RNA-seq analysis of prostate cancer cells [III]

city: Guangzhou

first: Xin

last: Chen

address: No. 100 Waihuan Xi Road, GuangzhouHigher Education Mega Center, Panyu District

institute: Guangdong University of Technology

summary: Androgen receptor (AR) plays key roles both in development of normal prostate gland and at all stages of prostate cancer. Due to the diversity of cell response to androgen stimulation, it’s necessary to investigate AR regulatory network at single cell level to fully understand how AR regulates transcription in prostate cancer cells. Here, we performed anti-androgen drug treated single cell RNA-seq profiling and transcriptome analysis on prostate cancer cell lines following 5α-dihydrotestosterone (DHT) stimulation. We sequenced 132 single cell transcriptomes in total and found gene expression heterogeneity among each cell. Finally, novel biological processes have been discovered combined with clinical data. This study provides a comprehensive profile to dissect hormone regulation in prostate cancer at single cell resolution.

overall_design: Single-cell RNA sequencing of prostate cancer cell VCaP.

first: Linging

last: Hu

first: Edwin

last: Cheung


title: Single-Cell Analysis Reveals Androgen Receptor Regulates ER-to-Golgi Trafficking  with CREB3L2 To Drive Prostate Cancer Progression

summary: Androgen receptor (AR) plays an essential role in normal prostate development and prostate cancer (PCa) progression. To understand the role of AR at the single-cell level, we performed single-cell transcriptome analysis on PCa cells stimulated with androgen and antiandrogen to reconstruct the dynamic and direct AR transcriptional network. Our work reveals that androgen stimulates the ER and Golgi stress response , promoting secreting protein trafficking, and inhibiting cell apoptosis. Moreover, we identify an ER-to-Golgi protein vesicle-mediated transport gene signature essential for maximal androgen-mediated ER-Golgi trafficking, cell proliferation, and association with PCa prognosis and progression. Notably, we show that AR collaborates with CREB3L2, XXX, to coordinately promote ER-Golgi trafficking of Golgi enzyme Mannosidase II  and PCa cell survival. Finally, we show the inhibition of the ER-Golgi transport process with Brefeldin A leads to tumor regression. Our study collectively reveals the heterogeneity  of PCa cell transcriptional response to androgen stimulation, demonstrates a functional role for increased ER-Golgi trafficking process, and provides a mechanism for how the process is augmented in PCa as well as the potential of targeting may provide novel treatment strategies.

overall_design: VCaP cells were treated with either ethanol (vehicle) or 100nM DHT for 2h, before being harvested for ChIP assay.

first: Lingling

first: Nitin

last: Narwade


title: Single-cell RNA-seq analysis of prostate cancer cells [II]

summary: Androgen receptor (AR) plays key roles both in development of normal prostate gland and at all stages of prostate cancer. Due to the diversity of cell response to androgen stimulation, it’s necessary to investigate AR regulatory network at single cell level to fully understand how AR regulates transcription in prostate cancer cells. Here, we performed normal growing and time-series 5α-dihydrotestosterone (DHT) stimulated single cell RNA-seq profiling and transcriptome analysis on prostate cancer cell lines. We sequenced 186 single cell transcriptomes in total and found gene expression heterogeneity among each cell. Finally, novel biological processes have been discovered combined with clinical data. This study provides a comprehensive profile to dissect hormone regulation in prostate cancer at single cell resolution.

overall_design: Single-cell RNA and bulk sequencing of normal growing VCaP and LNCaP, time course LNCaP cells stimulated by DHT.


title: Single-cell RNA-seq analysis of prostate cancer cells

summary: This SuperSeries is composed of the SubSeries listed below.


title: An unbalanced BMP-biomechanical niche reveals the plasticity of the mammalian myogenic lineage

city: Berlin

first: Xiaoyan

last: Wei

address: Thielallee 63

institute: Free University of Berlin

summary: Transcriptome analysis of muscle tissue from E15.5 fore limbs of wild type and Noggin-deficient mouse embryos indicates aberrant niche environment

overall_design: Muscle tissue of  E15.5 fore limbs of wild type and Noggin-deficient mouse embryos was collected and subjected to RNA extraction used for sequencing.

first: Arunima

last: Murgai

first: Sigmar

last: Stricker


title: A Population of M2 Macrophages Associated with Bone Formation

status: Public on Oct 09 2021

city: Houston

first: Alan

last: Davis

email: adavis3269@gmail.com, ardavis@bcm.edu

state: TX

address: One Baylor Plaza

department: Center for Cell and Gene Therapy

institute: Baylor College of Medicine

summary:
Purpose: We had previously published that very early during bone formation we observed a very transient cell that had the phenotype of brown fat and was important in creating a hypoxic microenvironment. This is critical for cartilage formation, the first stage in bone formation.
The purpose of the experiments in this manuscript were to determine the phenotype of these "brown fat" cells in more detail.
Methods: To determine the phenotype of these cells they were isolated by FACS for the beta 3 adrenergic receptor that we had previously found was a good marker for these cells during bone formation. After isolation, the cells both from HO and from the callus of a fracture were subjected to single cell RNA seq.
Results: The results of the single cell RNA seq showed that the cells in HO had many of the phenotypic markers of type 2 macrophages. However, they also retained the markers of brown fat including Ucp1 and Ucp2 as well as many markers of adipogenesis and extreme elevation of mitochondrial proteins. This mixture of proteins is required to burn oxygen in the microenvionment to create hypoxia. The cells made during fracture repair were further along in bone formation because they had other blood cells (B and NK cells) besides these macrophages. However, a small fraction was the same as the macrophages found during HO.
Conclusions: The type 2 macrophages made during bone formation had many of the properties of type 2 macrophages in the blood. However, they were also different from macrophages found in blood in that they lacked many of the key markers of blood macrophages. These unique macrophages are made rapidly at the start of bone formation and then disappear four days later probably differentiating to osteoclasts. However, they are critical in determining where the new bone forms as well as its size and shape.

overall_design: scRNA seq in both Heterotopic ossification (HO) and fracture repair. HO and callus were each bar-coded at the single-cell level, which were then combined before preparing the RNA.

first: Elizabeth

last: Olmsted-Davis


title: Anode respiration-dependent biological nitrogen fixation by Geobacter sulfurreducens

city: Fuzhou

first: jing

middle: xian

last: yue

address: No.15 Shangxiadian Road

institute: Fujian Agriculture and Forestry University

summary: Our study revealed a synergistic effect between biological nitrogen fixation and current generation by G. sulfurreducens, providing a green nitrogen fixation alternative through shifting the nitrogen fixation field from energy consumption to energy production and having implications for N-deficient wastewater treatment.

overall_design: Examination of the influence of NH4Cl on the cells. Cells cultured with NH4Cl addition was seted as control group.

first: Xianyue

last: Jing

first: Xing

last: Liu

first: Zhishuai

last: Zhang

last: Wang

first: Christopher

last: Rensing

first: Shungui

last: Zhou


title: Microbiota orchestrates monocyte reprogramming of the tumor microenvironment via type I IFN-STING axis to promote anticancer immunity

city: Bethesda

first: Maxwell

last: Lee

state: MD

address: Building 37, Room 5120

department: CCR

institute: National Cancer Institute, NIH

summary: This study sets out to investigate the mechanisms by which gut microbiota controls the mononuclear phagocyte compartment of the tumor microenvironment to elicit effective anti-tumor responses. This dataset contains single-cell RNA sequencing data of leukocytes from murine subcutaneous EL4 lymphomas from mice with intact or absent microbiota.

overall_design: EL4 tumor-infiltrating leukocytes sorted as LiveCD45+CD3- (CD3- leukocytes) or LiveCD45+CD3-CD19-NK1.1-TCRb-TCRgd-Ter119- (myeloid) from specific-pathogen-free (SPF) or germ-free (GF) mice and single-cell RNA-sequencing was performed on the 10X platform.

first: Khiem

last: Lam

first: Romina

last: Araya

middle: P

last: Goldszmid


title: Differentially expressed miRNA in recurrent implantation failure (RIF)

city: Jinan

first: Qian

address: JingLiu Road

department: Center for Reproductive Medicine

institute: Shandong Provincial Hospital Affiliated to Shandong University

summary: we utilized Exiqon miRCURY™ LNA Array (v18.0, Exiqon, Vedbaek, Denmark)  to establish miRNA expression profiles in the endometrial tissues at the time of embryo implantation(day 7 after ovulation) from 8 RIF patients and 10 matched controls. A total of 157 miRNAs exhibited distinct expression patterns in RIF patients as opposed to controls (fold change >2.0 and P-value <0.05).

platform_organism:
Homo sapiens
Mus musculus
Rattus norvegicus

overall_design: Our study was designed to demonstrat altered miRNA profiles in the endometrial tissues from RIF patietns. Functional studies will be further confirmed. This study would provide new insights into epigenetic pathogenesis for RIF, raising the prospect that modulating disordered miRNAs might be helpful for improving implantation rates in reproductive treatment.

first: Junhao

last: Yan


title: Expression data from prostate cancer LNCaP cells in response to Ev or ID1 plasmid overexpression

city: Portland

first: David

last: Qian

address: 3303 SW Bond Ave, CH14R

institute: Oregon Health & Science University

summary:
ID1 is a negative transcription regulator, its effects on gene expression in regular or gypoxic conditions are investigated
We used gene expression array to determine the globe gene expression changes due to Ev or ID1-overexpression in 20% or 1% oxygen

overall_design: human prostate cancer cell lines with stable transfection of Ev or ID1 plasmids were cultured in regular 20% oxygen or hypoxic 1% oxygen conditions for 24 hours

middle: Z


title: Sex-Specific Microglia State in the Neuroligin-4 Knock-Out Mouse Model of Autism Spectrum Disorder

first: Dilansu

last: Guneykaya

address: 220 Longwood Avenue

department: Neurobiology

institute: Harvard Medical School

summary: Neuroligin-4 (NL4) loss-of-function mutations are strongly associated with monogenic heritable abnormalities linked with Autism Spectrum Disorder (ASD). NL4 mutation in mice causes ASD related alterations in both synaptic and behavioral phenotypes. Since microglia closely regulate synaptic development and are implicated as key players in ASD development and progression, we here studied microglial properties in the NL4-knock-out (NL4-/-) mouse model. We show that loss of NL4 caused altered behavior and impaired hippocampal gamma oscillations predominantly in male mice. In parallel, microglial density, morphology, and response to injury specifically in the CA3 region of the hippocampus were altered in NL4-/- males only. A transcriptomic and proteomic analysis revealed strong sexual dimorphism on molecular alterations in microglia of NL4-/-. Together, these results indicate that loss of NL4 affects not only neuronal network activity and behavior, but also changes the phenotype of microglia in a sex by genotype interaction .

overall_design: 2 Biological replicates were analyzed for controls and 5 biological replicates per group were analyzed for phagocytosis group.

first: Susanne

last: Wolf

first: Bilge

last: Ugursu


title: Cas12a variants designed for lower genome-wide off-target effect through stringent PAM recognition

city: Wuhan

first: chen

last: peng

address: College of Life Sciences, Wuhan University

department: College of Life Sciences

institute: Wuhan University

summary: our findings show that the Cas proteins could be designed to be stringent in PAM recognition to reduce their off-target effects , which points out a promising and practical way for the minimization of the off-targeting in genome editing.

overall_design: To minimize off-targeting for gene editing, here we engineered a variant of LbCas12a (termed Lb-K538R) with more stringent PAM recognition, lower off-targeting, and similar editing efficiency in vivo compared with LbCas12a. We also demonstrate that Lb2Cas12a from Lachnospiraceae bacterium MA2020 exhibits extensive gene-editing activities in mammalian cells. Similar to Lb-K538R, the designed Lb2Cas12a variant (termed Lb2-K518R) also has lower off-target effects and is more stringent in PAM recognition from YYN to be TYN (Y is T or C, N is A, T, C, or G), which supplies more possible target site selections with low off-targeting as compared with the common TTTV (V is A, G, or C) PAM.

first: Lei

last: Yin


title: Unveiling E2F4, TEAD1 and AP-1 as regulatory transcription factors of the replicative senescence program by multiomics analysis

status: Public on Oct 08 2021

city: Beijing

first: liping

last: liu

state: Beijing

address: Yiheyuanlu 5#

institute: Peking University

summary: Senescence, a stable state of growth arrest, affects many physiological and pathophysiological processes, especially aging. Previous work has indicated that transcription factors (TFs) play a role in regulating senescence. However, a systematic study of regulatory TFs during replicative senescence (RS) using multi-omics analysis is still lacking. Here, we generated time-resolved RNA-seq, reduced representation bisulfite sequencing (RRBS) and ATAC-seq datasets during RS of mouse skin fibroblasts, which demonstrated that an enhanced inflammatory response and reduced proliferative capacity were the main characteristics of RS in both the transcriptome and epigenome. Through integrative analysis and genetic manipulation, we found that transcription factors E2F4, TEAD1 and AP-1 are key regulators of RS. Overexpression of E2f4 improved cellular proliferative capacity, attenuated SA-β-Gal activity and changed RS-associated differentially methylated sites (DMSs). Moreover, knockdown of Tead1 attenuated SA-β-Gal activity and partially altered the RS-associated transcriptome. In addition, knockdown of Atf3, one member of AP-1 superfamily TFs, reduced Cdkn2a (P16) expression in pre-senescent fibroblasts. Taken together, results of this study identified transcription factors regulating the senescence program through multi-omics analysis, providing potential therapeutic targets for anti-aging.

overall_design: Cut&Tag profile of P2 and P9 mouse fibroblasts, targeting at E2F4 and ATF3

first: Yuting

first: Liping

first: Yifan

last: Song


title: Phage-Based Profiling of  Rare Single Cells Using Nanoparticle-Directed Capture

city: Toronto

first: Yuan

last: Ma

address: 144 College St.

institute: University of Toronto

summary: To extend the applicability of our approach, we tested a more extensively multiplexed pool of phages which contained ~50 clones with each phage equally distributed across the pool. Most of the factors tested are associated with cancer progression.

overall_design: Here we demonstrate an integrated microfluidic chip that uses magnetic nanoparticles to capture single tumor cells with high efficiency, permits on-chip incubation and facilitates in situ cell-surface protein expression analysis. Combined with phage-based barcoding and next-generation sequencing technology, we were able to monitor changes in the expression of multiple surface markers stimulated in response to CTC adherence.

first: Kangfu

last: Chen

first: Fan

last: Xia

first: Randy

last: Atwal

first: Hansen

first: Sharif

last: Ahmed

first: Lia

last: Cardarelli

first: Irene

last: Lui

first: Bill

last: Duong

first: Zongjie

first: James

last: Wells

first: Sachdev

last: Sidhu

first: Shana

last: Kelley


title: Immunomodulatory effects of ponesimod on human astrocytes

city: La Jolla

first: Yasuyuki

last: Kihara

state: California

address: 10901 N Torrey Pines Rd

institute: Sanford Burnham Prebys Medical Discovery Institute

summary: Ponesimod is a slective S1PR1 antagonist and is approved for the treatment of Multiple Sclerosis by FDA. To determine how ponesimod controls neuroinflammatory responses in astrocytes, single-cell RNA-sequencing (scRNA-seq) was conducted in cytokine mixture-stimulated human primary astrocytes in the presence or absence of ponesimod.

overall_design: Human primary astrocytes (Lonza; Cat# CC-2565) were stimulated with or without cytokine mixture (25 ng/mL each; TNF-a, IFN-g, IL-17, IL-1b, and IL-6) in the presence or absence of ponesimod for 24 h or 48 hrs. Single-cell capture (target capture of 10,000 cells per sample) and library preparation was conducted using Chromium Single Cell 3' GEM, Library & Gel Bead Kit v3 (10x Genomics, Cat # PN-1000075) and Chromium Chip B Single Cell Kit (10X Genomics, Cat # PN-1000074) and according to manufacturer's instruction. Single-nucleus libraries were sequenced on the Illumina NovaSeq 6000 machine at GENEWIZ, Inc. (U.S.)  Raw FASTQ files were input into the cellranger count pipeline (Cell Ranger V4.0.0, 10x Genomics) to align reads to the GRCh38 human genome.

first: Kihara

last: Yasuyuki

first: Zhu

last: Yunjiao


title: Transcriptome analysis of 1-year-old Panax vietnamensis Ha et Grushv.

city: Hà Nội

first: Ly

middle: Hàn

last: Lưu

state: Cau giay District

address: Trần Đăng Ninh, Cầu Giấy

country: Viet Nam

institute: Institute of Genome Research

summary:
Purpose: Panax vietnamensis Ha et Grushv., which contains various valuable ginsenosides, is an important herbal medicine of Vietnam. However, it is an endangered species listed in Vietnam Red Data Book due to over-harvseting. Investigation about genomic or trancriptomic resources is one of the necessary activities to conserve P. vietnamensis
Materials and Methods:. In this study, we sequenced the transcriptomes of 1-year-old P. vietnamensis from leaves and roots using Illumina NovaSeqTM6000 system.
Results:  A total of 60,254,062 and 64,588,528 reads was obtained and then assembled into 45,495 and 49,133 unigenes for leaves and roots, respectively. More than 60% unigenes from two organs  were functional annotated using at least one database among Kyoto Encyclopedia of Genes and Genomes, Pfam, Gene Ontology, NCBI non-redundant Protein, and Evolutionary genealogy of genes. Further, the predominant transcripts of each cDNA library were analyzed for different gene expression identification. In addition, 457 unigenes encoding enzymes involved in triterpenoid saponin biosynthesis via the mavelonate (MVA) and the non-MVA (also named as MEP pathways) were discovered.

overall_design: mRNA profiles of 1-year-old P.vietnamensis leaves and roots

first: Hien

middle: T

last: Le

middle: H

last: Luu

first: Linh

middle: N

last: Nguyen

first: Nam

last: Vu

first: Hang

middle: L

last: Pham


title: Identification of the downstream target genes of miR-138 in gastric cancer

city: Shiyan

first: Qin

last: Shanshan

address: Renmin Road 30, Maojian District

institute: Institute of Basic Medical Sciences, School of Basic Medical Sciences, Hubei University of Medicine

summary: Our data showed that EIF4EBP1, NINJ1, CLIC3, STMN3, SPINK4, CD44, GSN, PRSS23 et al. were strongly downregulated by miR-138-mimics in gastric cancer (GC) cell lines. Besides, miR-138-5p has a profound effect on EMT signaling and proliferation signaling. In the EMT signaling, miR-138-5p negatively regulates the expression of SOX4, VIM, TGFB1, BMP1, ECM1 and CXCL8; In the cell proliferation signaling, miR-138-5p could negatively regulates the expression of PCNA, RHOC and CCND3.

overall_design: To reveal the possible target genes of miR-138 in gastric cancer, we transfected miR-138-5p mimics in GC cells. The potent miR-138 target genes were identified by deep sequencing, using Illumina HiSeq 4000.

first: Shanshan

last: Qin

first: Dandan

last: Li


title: Gene expression analysis under normal and stress conditions during in-vitro embryo development and subanalysis of gene expression in the blastocyst stage according to its Mitoscore®.

city: Valencia

first: Mª José

last: de los Santos Molina

address: C/ Plaça de la Policia Local, 3

country: Spain

institute: IVIRMA Valencia

summary: We analyzed the differences in the gene expression profile, especially pathways related to mitochondrial activity, between human embryos of different developmental stages, different Mitoscore® values and under stress conditions such as embryo arrest and the vitrification/warming process. In total, 44 vitrified specimens were collected for RNA-seq analysis, including 11 MII oocytes, 10 non-arrested cleavage-stage embryos, 5 arrested cleavage-stage embryos and 18 blastocysts (10 blastocysts cultured for 4-5 hours after warming and 8 blastocysts cultured for 0 hours after warming). All specimens were warmed and whole sampled in PCR tubes with 2 μL of 10× Reaction Buffer (SMART-Seq v4 Ultra-Low Input RNA kit for Sequencing, Takara Bio, USA). Amplified cDNA obtained from mRNA was purified using AMPure XP magnetic beads (Illumina, CA, USA). Libraries were constructed using NexteraXT DNA sample preparation (Illumina, CA, USA) and sequencing was done in duplicate in a total of four runs using an Illumina NextSeq 500 (Illumina, CA, USA) with a 300-nucleotide read length in a paired-end design (150-bp fragments). FastQC was used for checking the quality of the raw sequence data. Those fragments that did not meet quality requirements were trimmed using Trimmomatic. Alignment and quantification were performed using the Salmon algorithm (reference genome GRCh38). We observed that disruption of normal in-vitro culture and high Mitoscore® values are related to an upregulation of cellular stress pathways in human embryos. Despite the crucial role of mitochondria in cellullar stress, an upregulation of mitochondrial activity pathways is only observed during embryo arrest.

overall_design: Analysis of general and mitochondrial-related differences in gene expression between different embryo stages and conditions: normal in-vitro culture (MII oocytes> non-arrested cleavage-stage embryos> blastocysts), embryo arrest (arrested cleavage-stage embryos vs non-arrested cleavage-stage embryos), vitrification/warming recovery (blastocysts cultured for 4-5 hours after warming vs blastocysts cultured for 0 hours after warming)  and blastocysts Mitoscore® level (high Mitoscore® blastocysts vs low Mitoscore® blastocysts).

first: delosSantos

last: María José

first: Pérez-Sánchez

last: Marta


title: RNA sequencing of gene expression in fibroblasts derived from idiopathic pulmonary fibrosis and control lungs

city: Perth

first: Zhenjun

last: Deng

state: Western Australia

address: 35 Stirling Highway

country: Australia

institute: The University of Western Australia

summary: RNA sequencing has been performed to investigate the transcriptomic profile of fibroblasts derived from apical and basal regions of idiopathic pulmonary fibrosis (IPF) and control (CTR) lungs. Lung fibroblasts were cultured onto T25 flasks until full confluence was reached. Fibroblasts were then harvested by trypsinization using 0.05% trypsin-EDTA and washed in Phosphate Buffered Saline (PBS). RNA was extracted using the RNeasy Mini kit protocol. Samples were then sent to the Australian Genome Research Facility (AGRF) where 1μg of RNA was submitted for next-generation sequencing. Once raw data returned, bioinformatics analysis was conducted. Results revealed little difference at the transcriptomic level between apical lung fibroblasts isolated from IPF and CTR donors. In contrast, there was a significant difference in gene expression between IPF and CTR lung basal fibroblasts with 90 differentially expressed genes (DEGs) identified in IPF basal fibroblasts. Gene ontology analysis of these 90 DEGs suggested that the most important functions were associated with receptor ligand activity, cell adhesion molecule binding, and integrin binding. Furthermore, fibroblasts isolated from the basal and apical sites of control lung were not significantly different. Interestingly, the lung basal fibroblasts from IPF patients were significantly different in their transcriptomic profile to that of apical fibroblasts from the same patients. 303 DEGs were identified in IPF basal fibroblasts compared with IPF apical fibroblasts. According to the GO analysis on these 303 DEGs, the most important function of the identified dysregulated genes was associated with the composition and function of the extracellular (ECM). Further pathway analysis identified 4 signaling pathways, namely arrhythmogenic right ventricular cardiomyopathy, calcium signaling pathway, dilated cardiomyopathy, and hematopoietic cell lineage. An interesting finding was that these IPF basal fibroblasts were clustered into two groups (Group 1 and Group 2). Analysis on these two groups found 3594 DEGs in Group 1 compared to Group 2. Go analysis suggested most of the identified dysregulated genes was involved in the composition and function of the ECM. Pathway analysis identified 28 dysregulated signaling pathways, including PI3K-Akt signaling pathway, TGF-β signaling pathway and Wnt signaling pathway. The results of the transcriptome analysis intimate that the IPF is more predominant in the basal lungs and the fibroblast-derived from the basal region of the fibrotic lungs may serve as a central role in IPF. Moreover, the dramatic difference between two groups of IPF basal fibroblasts might be linked to the differential response to the same drug occurred in IPF patients.

overall_design: total RNA profiles of fibroblasts derived from apical and basal regions of idiopathic pulmonary fibrosis (IPF) and control (CTR) lungs

first: Tylah

last: Miles


title: p53-Dependent Induction of P2X7 on Hematopoietic Stem and Progenitor Cells Regulates Hematopoietic Response to Genotoxic Stress [RNA-seq]

city: Montreal

first: Anastasia

last: Nijnik

state: Quebec

address: Rm 368 - 3649 Promenade Sir William Osler

department: Physiology

institute: McGill University

summary: Stem and progenitor cells are the main mediators of tissue renewal and repair, both under homeostatic conditions and in the response to physiological stress and injury. Hematopoietic system is responsible for the regeneration of blood and immune cells, and is maintained by bone marrow resident hematopoietic stem and progenitor cells (HSPCs). Hematopoietic system is particularly susceptible to injury in response to genotoxic stress, resulting in risk of bone marrow failure and secondary malignancies in cancer patients undergoing radiotherapy. Here we analyze the in vivo transcriptional response of HSPCs to genotoxic stress in a mouse whole body irradiation model, and together with p53 ChIP-Seq and studies in p53-knockout (p53KO) mice, characterize the p53-dependent and p53-independent branches of this transcriptional response. Our work demonstrates the p53-independent induction of inflammatory transcriptional signatures in HSPCs in response to genotoxic stress, and identifies multiple novel p53-target genes induced in HSPCs in response to whole body irradiation. In particular, we establish the direct p53-mediated induction of P2X7 expression on HSCs and HSPCs in response to genotoxic stress. We further demonstrate the role of P2X7 in hematopoietic response to acute genotoxic stress, with P2X7-deficiency significantly extending mouse survival in irradiation-induced hematopoietic failure. We also demonstrate the role of P2X7 in the context of long-term HSC regenerative fitness following sublethal irradiation. Overall our studies provide important insights into the mechanisms of HSC response to genotoxic stress and further suggests P2X7 as a target for pharmacological modulation of HSC fitness and hematopoietic response to genotoxic injury.

overall_design: We performed RNA-Seq transcriptional profiling on FACS-sorted Lin-cKit+Sca1+CD150+ HSPCs from wild type (WT) mice and Tp53-knockout (p53KO) mice (JAX 002101), untreated and at 3 hours after 3Gy whole body irradiation (IR). RNA was isolated using the MagMAX total RNA kit (Ambion, Life Technologies), and quality assessed using Bioanalyzer RNA Pico chips (Agilent). rRNA depletion and library preparation were performed using the SMARTer Stranded RNA-Seq kit (Takara Clontech). The libraries were sequenced on Illumina HiSeq 2000 and 2500 sequencers in paired-end 50bp configuration (two sequencing runs).

first: HanChen

last: Wang

first: Jessica

last: Petrov

first: Jad

last: Belle

first: David

last: Langlais


title: Introgression of genes and mechanisms  conferring tolerance to iron toxicity from wild rice species Oryza meridionalis into domesticated rice Oryza sativa

city: Porto Alegre

first: Andriele

last: Wairich

state: Rio Grande do Sul

address: Av. Bento Gonçalves, 9500

department: Center for Biotechnology

country: Brazil

institute: Universidade Federal do Rio Grande do Sul

summary: Iron toxicity is one of the most common mineral disorders affecting Oryza sativa production in flooded lowland fields. Efforts have been made to develop new rice varieties tolerant to Fe toxicity (+Fe). Oryza meridionalis is an endemic from Northern Australia and grows in regions with Fe rich soils, which may provide Fe tolerance genes and mechanisms that can be used for adaptive breeding. Aiming to understand tolerance mechanisms in rice, we screened a population of interspecific introgression lines (IL) from a cross between O. sativa and O. meridionalis for the identification of QTLs contributing to Fe excess tolerance. Six putative QTLs were identified. A line carrying one introgression from O. meridionalis on chromosome 9 associated with one QTL for leaf bronzing score was identified as tolerant in terms of lipid peroxidation and electrolyte leakage despite presenting very high shoot Fe concentrations. Further physiological, biochemical, ionomic and transcriptomic analyses showed that the IL tolerance could be partly explained by Fe partitioning between the leaf sheath and culm. After the in silico construction of an interspecific hybrid genome to map the sequences from transcriptomic analysis, we identified 47 and 27 genes from O. meridionalis up and down-regulated, respectively, by Fe treatment on the tolerant IL. Among possible genes associated with shoot-based tolerance, we identified metallothionein-like proteins, genes from glutathione S-transferase family and transporters from ABC and Major Facilitator Superfamily. This is the first work to demonstrate that introgressions of O. meridionalis in O. sativa genome confer increased tolerance to +Fe

sample_organism: Oryza sativa Japonica Group x Oryza meridionalis

overall_design: We analyzed the shoot transcriptomic response to Fe excess in two species from the Oryza genus: Oryza sativa (cv. Curinga) and Oryza meridionalis (WG 2112), and in one introgression line from a cross between these two species, CM23. Plants were grown in greenhouse conditions under control and Fe toxicity treatments for three weeks and then exposed to treatment. Three biological replicates were collected after seven days of treatment. Total RNA was extracted from shoots and libraries were generated and sequenced at BGI Genomics Co. using HiSeq Illumina 4000.

first: Ben

last: Oliveira

first: Lin-Bo

last: Wu

first: Varunseelan

last: Murugaiyan

first: Marcia

last: Margis-Pinheiro

first: Janette

last: Fett

first: Felipe

middle: K

last: Ricachenevsky

first: Michael

last: Frei


title: p53-Dependent Induction of P2X7 on Hematopoietic Stem and Progenitor Cells Regulates Hematopoietic Response to Genotoxic Stress

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: p53-Dependent Induction of P2X7 on Hematopoietic Stem and Progenitor Cells Regulates Hematopoietic Response to Genotoxic Stress [ChIP-seq]

overall_design: To map p53 DNA binding sites, p53 chromatin immunoprecipitation sequencing (ChIP-Seq) was performed in multipotent progenitor cells HPC7 and B cell progenitors Ba/F3, both selected for their p53 wild type status. In parallel, ChIP-Seq was done in Ba/F3s following irradiation (3Gy, 3 hours).


title: RNAseq analyses of A549 cells treated with combinations of interferon-gamma and a novel anti-cancer reagent AX0085

city: Seoul

first: Jaesang

last: Kim

address: 52, Ewhayeodae-gil

country: South Korea

institute: Ewha Womans Universiyt

summary: A549 cells were used to examine the effect of AX0085, a novel kinase inhibitor. In particular, its effect in reversing the effect of interferon gamma was determined.  Combination treatment of two reagents were to be  compared with no treatment control and single treatment of either AX0085 or interferon gamma.

overall_design: Duplicate samples of A549 cells under 4 different condtions are prepared: Np treatment, interferon gamma treatment, AX0085 treatment, & interferon gamma+ AX0085 treatment.

first: Jusong

first: Haeyeon

last: Jang

first: Gyu

last: Lee


title: FBXO32 is a KIT-downstream prosurvival factor in GIST

city: Mainz

first: Sergi

last: Sayols

address: Ackermannweg 4

institute: Institute of Molecular Biology, Mainz

summary: GISTs are the most common mesenchymal tumors and they display neuromuscular features. Gain-of-function mutations in KIT or PDGFRA are the initiating event in most of the cases and KIT oncogenic signaling is essential throughout all the disease. Interestingly, KIT oncogenic signaling is driven by PI3K/mTOR and RAS/MAPK pathways regardless of KIT primary or secondary mutations. Hence, dissection of KIT-downstream pathways may result in the identification of novel KIT critical effectors and a new opportunity to overcome disease heterogeneity of resistance mechanisms in GIST.  Using human clinically representative GIST cell models, we undertook pharmacological and functional screenings such as Cell titer glo, kinase activation, apoptosis induction and proliferation to identify key signaling nodes within PI3K/mTOR and RAS/MAPK pathways. In vitro and in vivo validations were undertaken to model novel therapeutic strategies. Transcriptomic analysis (RNAseq) found essential genes co-regulated by both pathways. Functional studies, such as immunofluorescence, kinase activation, flow cytometry and proliferation were performed on a viral gene overexpression and knock-down models to elucidate their regulation and potential role in GIST biology.  PI3K/mTOR and MEK1/2 are the most essential KIT-downstream mediators. Single node inhibition did not yield sustained antiproliferative and apoptotic effect, but simultaneous intermittent ablation was synergistic in vitro and in vivo, supporting the critical role of these two pathways.Transcriptomic analyses underscored FBXO32 (a SCF E3 ubiquitin-ligase and the main effector of muscular atrophy) as the most differentially expressed gene co-regulated by PI3K/mTOR and RAS/MAPK pathways. Functional studies demonstrated that FBXO32 is transcriptionally activated by FOXO3a which, in turn, is regulated by PI3K/mTOR and RAS/MAPK pathways. Notoriously, FBXO32 proved to have a pro-survival role in GIST cells. Microarray studies hinted a participation of FBXO32 in cell cycle arrest, which has been previously described as an adaptive pro-survival mechanism in GIST cells. This mechanism was further confirmed by functional studies. Remarkably, FBXO32 was expressed upon KIT inhibition in a panel of GIST cell lines, regardless the mutational status of KIT, and in a gene set database from GIST patients treated with first-line treatment, imatinib.  PI3K/mTOR and MEK1/2 are the critical mediators of KIT oncogenic signaling in GIST, and their intermittent co-inhibition is an effective and well-tolerated therapeutic strategy to treat GIST tumors, independently of their KIT mutational status. FBXO32 emerges as a common mediator of KIT-downstream PI3K/mTOR and RAS/MAPK pathways with a critical pro-survival role in GIST participating in cell cycle arrest as pro-survival mechanism.

overall_design: 4 different GIST cell lines (GIST-T1, GIST882, GIST-T1/670, GIST-T1/816) treated at 4 and 24 hours with: GSK-212 100nM, GDC-0980 500nM and the combination of both disolved in DMSO. All treatments were performed in serum-free media

first: Alfonso

last: Garcia-Valverde

first: Cesar

last: Serrano


title: Mulberry, an edible plant, bark derived exosomes-like nanoparticles (MBELN) treatment of B6 mice for 7 days

status: Public on Oct 07 2021

city: LOUISVILLE

first: Mukesh

last: Sriwastva

state: KY

address: 505 S Hancock St

institute: University of Louisville


title: Mulberry, an edible plant, bark derived exosomes-like nanoparticles (MBELN) treatment of B6 mice for 7 days [Colon]

summary:
MBELN isolated mulberry bark purified by sucrose gradient were orally gavaged to B6 mice for 7 days once in a day at aconcentration of 10 x 10E+09 particle/100 µl.
After 7 days, colon tissue were harvested. RNA was isolated and sent to invitorgen for Affymetrix array. The purpose of the experiment was to see which set of genes go up or down after MBELN treatment and their link to gut homeostasis.

overall_design: We used affymetric array to see differential gene expression after the MBELN treatment in B6 mice colon tissue.

last: Kumar Sriwastva

first: Yun

last: Teng

first: Huang-Ge

last: Zhang


title: Inhibition of vascular growth by modulation of the AEA/FAAH axis

city: Bochum

first: Daniela

last: Wenzel

address: Universitätsstr. 150

department: Department of Systems Physiology

institute: University of Bochum

summary: Inhibition of FAAH and AEA accumulation reduces angiogenesis.

overall_design: Total RNA obtained from HUVEC treated with URB (20 μM), AEA (10 μM) or ethanol compared to untreated HUVEC.


title: Single-cell RNA-seq analysis of prostate cancer cells [I]

city: Guangzhou

first: Xin

last: Chen

address: No. 100 Waihuan Xi Road, GuangzhouHigher Education Mega Center, Panyu District

institute: Guangdong University of Technology

summary: Androgen receptor (AR) plays key roles both in development of normal prostate gland and at all stages of prostate cancer. Due to the diversity of cell response to androgen stimulation, it’s necessary to investigate AR regulatory network at single cell level to fully understand how AR regulates transcription in prostate cancer cells. Here, we performed time-series and anti-androgen drug treated single cell RNA-seq profiling and transcriptome analysis on prostate cancer cell lines following 5α-dihydrotestosterone (DHT) stimulation. We sequenced 1861 single cell transcriptomes in total and found gene expression heterogeneity among each cell. Finally, novel biological processes have been discovered combined with clinical data. This study provides a comprehensive profile to dissect hormone regulation in prostate cancer at single cell resolution.

overall_design: Single-cell RNA sequencing of prostate cancer cell line VCaP.

first: Linging

last: Hu

first: Edwin

last: Cheung


title: Mulberry, an edible plant, bark derived exosomes-like nanoparticles (MBELN) treatment of B6 mice for 7 days [Crypt]

summary:
MBELN isolated mulberry bark purified by sucrose gradient were orally gavaged to B6 mice for 7 days once in a day at aconcentration of 10 x 10E+09 particle/100 µl.
After 7 days, colon specific crypt cells were isolated. RNA was isolated and sent to invitorgen for Affymetrix array. The purpose of the experiment was to see which set of genes go up or down after MBELN treatment and their link to gut homeostasis.

overall_design: We used affymetric array to see differential gene expression after the MBELN treatment in B6 mice colon specific crypt.


title: UVB drives transcriptomic and epigenetic reprogramming which can be protected by antioxidant sulforaphane in human skin keratinocytes [Methyl-seq]

city: Piscataway

first: Hsiao-Chen Dina

last: Kuo

address: 160 frelinghuysen road

department: Ernest Mario School of Pharmacy

institute: Rutgers University

type: Methylation profiling by high throughput sequencing

summary: Continuous exposure to ultraviolet (UV) is one of the main factors contributing to skin carcinogenesis. Sulforaphane (SFN) is a potent antioxidative agent which has potential to prevent the UV-induced skin cell transformation. We characterized the transcriptome and CpG methylation profile of human keratinocytes (HaCaT cells) treated with UVB and/or SFN using RNA sequencing.

overall_design: Samples of HaCaT cells were used for extraction of DNA and RNA; DNA was enriched by Agilent SureSelect Kit then bisulfite converted methyl-seq was performed. RNA-seq was performed with routine. Strategy. This dataset contains data for the Methyl-seq only; Check the other SubSeries under this SuperSeries for RNA-seq data.

first: Hsiao-Chen

middle: D

first: Shanyi

last: Li

first: Lujing

last: Wang

first: Renyi

last: Wu

first: Davit

last: Sargsyan

first: Ah-Ng

last: Kong


title: Krüppel-Like Factor 5 Regulates Airway Epithelial Cell Identity and Function [RNA-seq]

city: Cleveland

first: Ann

last: Harris

state: Ohio

address: 2109 Adelbert Rd Building

department: Genetics and Genome Sciences

institute: Case Western Reserve University

summary: A complex network of transcription factors regulates genes involved in establishing and maintaining key biological properties of the human airway epithelium. Here we characterize the role of Krüppel-Like Factor 5 (KLF5), in controlling essential pathways of epithelial cell identity and function in the human lung. RNA-seq following siRNA-mediated depletion of KLF5 in the Calu-3 lung epithelial cell line identified significant enrichment of genes encoding chemokines and cytokines involved in the proinflammatory response, and also components of the junctional complexes mediating cell adhesion. To determine direct gene targets of KLF5, we defined the cistrome of KLF5 using ChIP-seq in both Calu-3 and 16HBE14o- lung epithelial cells lines. Occupancy site concordance analysis revealed that KLF5 co-localized with the active histone modification H3K27ac and also with binding sites for CCAAT Enhancer Binding Protein Beta (C/EBPβ). Depletion of KLF5 increased both the expression and secretion of important chemokines, a response that was enhanced following exposure to Pseudomonas aeruginosa lipopolysaccharide. Wound scratch assays in Calu-3 cells exhibited faster rates of repair after KLF5 depletion than did negative controls. Similarly, CRISPR-mediated KLF5-null 16HBE14o- cells also showed enhanced wound closure.  These data reveal a pivotal role for KLF5 in coordinating epithelial functions relevant to human lung disease.

overall_design: 6 RNA-seq (single end) datasets, 3 replicates between 2 treatment groups

first: Alekh

last: Paranjapye

first: Monali

last: NandyMazumdar

first: James

last: Browne

first: Shih-Hsing

last: Leir


title: Genome-wide maps of chromatin state and RNA-seq in WT and Dnmt3a KO BMDMs (bone marrow derived macrophages)

city: Boston

first: Peter

middle: Geon

last: Kim

address: 360 Longwood Ave, Lc8202

department: Hematology/Oncology

institute: Dana Farber Cancer Institute


title: In planta dynamics, transport-biases and endogenous functions of mobile siRNAs in Arabidopsis

city: Zürich

first: Alexis

last: Sarazin

address: Universitätstrasse 2

country: Switzerland

institute: ETH

type: Non-coding RNA profiling by high throughput sequencing

summary:
In  RNA  interference  (RNAi),  small-interfering  (si)RNAs  processed  from  double-stranded RNA guide ARGONAUTE(AGO) proteins to silence sequence-complementary  RNA/DNA.  Plant  RNAi  can  propagate  locally  and  systemically,  but despite recent mechanistic advances, basic questions/hurdles remain unaddressed. For  instance,  RNAi  is  inferred  to  diffuse  through  plasmodesmata,  yet  how  its dynamics  in  planta  compares  with  that  of  established  symplastic-diffusion  markers remains  unknown.  Also  unknown  is  why  select  siRNA  species,  or  size-classes thereof,  are  recovered  in  RNAi-recipient tissues,  yet  only  under  some  experimental settings. Finally, RNAi shootward movement in micro-grafted Arabidopsis necessary to study its presumptive transgenerational effects– has not been achieved thus far and endogenous functions of mobile RNAi remain scarcely documented.

Focusing  on  non-amplified  RNAi  in  Arabidopsis,  we  show  here  that  (i)  transgenic RNAi-movement, although symplasmic, only partially  recapitulates the diffusion pattern  of  free GFP  in  planta, (ii) the  presence/absence  of  specific  AGOs in incipient/traversed/recipient  tissues  likely  explains  the  apparent  siRNA-selectivity observed  during  vascular  movement,  (iii)  stress  application  allows  endo-siRNA translocation  against  the  shoot-to-root  phloem  flow,  and  (iv)  mobile  endo-siRNAs generated  from  a  single  inverted-repeat(IR)  locus,  have  the  potential  to  regulate hundreds of transcripts.

overall_design: 2 samples examined: shoot apex from Arabidopsis thaliana Col0 wild type and ir71 mutant

first: Christopher

last: Brosnan

first: Emanuel

last: Devers

first: Peiqi

last: Lim

first: Satu

last: Lehesranta

first: Ykä

last: Helariutta

first: Olivier

last: Voinnet


title: IFN-α kinoid in systemic lupus erythematosus: Affymetrix Data

city: Brussels

first: Frédéric

last: HOUSSIAU

address: Avenue Hippocrate 10

department: Institut de recherche expérimentale et clinique (IREC)

country: Belgium

institute: Université catholique de Louvain

summary:
Objective: To evaluate the efficacy and safety of the immunotherapeutic vaccine interferon-α kinoid (IFN-K) in a 36-week (W) phase IIb, randomised, double-blind, placebo (PBO)-controlled trial in adults with active systemic lupus erythematosus (SLE) despite standard of care.
Methods: Patients with SLE (185) with moderate to severe disease activity and positive interferon (IFN) gene signature were randomised to receive IFN-K or PBO intramuscular injections (days 0, 7 and 28 and W12 and W24). Coprimary endpoints at W36 were neutralisation of IFN gene signature and the BILAG-Based Composite Lupus Assessment (BICLA) modified by mandatory corticosteroid (CS) tapering.
Results: IFN-K induced neutralising anti-IFN-α2b serum antibodies in 91% of treated patients and reduce the IFN gene signature (p<0.0001). Modified BICLA responses at W36 did not statistically differ between IFN-K (41%) and PBO (34%). Trends on Systemic Lupus Erythematosus Responder Index-4, including steroid tapering at W36, favoured the IFN- K and became significant (p<0.05) in analyses restricted to patients who developed neutralising anti-IFN-α2b antibodies. Attainment of lupus low disease activity state (LLDAS) at W36 discriminated the two groups in favour of IFN-K (53% vs. 30%, p=0.0022). A significant CS sparing effect of IFN-K was observed from W28 onwards, with a 24% prednisone daily dose reduction at W36 in IFN-K compared with PBO (p=0.0097). The safety profile of IFN-K was acceptable.
Conclusions: IFN-K induced neutralising anti-IFN-α2b antibodies and significantly reduced the IFN gene signature with an acceptable safety profile. Although the clinical coprimary endpoint was not met, relevant secondary endpoints were achieved in the IFN-K group, including attainment of LLDAS and steroid tapering.
Trial registration number NCT02665364.

overall_design: 185 patients with active SLE with a positive IFN gene signature were randomised, with 93 assigned to PBO and 92 to IFN-K. At randomisation (D0) and W12, W24 and W36, the IFN gene signature, including the 21 IFN-regulated genes, was evaluated by Affymetrix. This was done on 177 patients, among which 169 have completed all visits until W36. RNA was extracted, quality assessed, labelled, hybridised to GeneChip human genome U133 Plus 2.0 arrays, washed, stained and scanned using GeneChip Scanner 3000 7G. Affymetrix CEL files were uploaded to Affymetrix Expression Console software V.1.4.1. Analysis and Robust multichip Analysis normalisation of raw data were performed per batch, and raw dataset was normalised at once.

first: Frederic

last: Houssiau

first: Aikaterini

last: Thanou

first: Thérèse

last: Croughs

first: Michael

last: Tee


title: Human tRF&tiRNA Sequencing for exosomes derived from ectopic endometrial tissues in EMs patients

city: Ningbo

first: yichen

last: chen

address: NO.46 Yangshan Road，Ningbo

institute: Ningbo Institute of Medical Science

summary: We digested and extracted tissue exosomes from fresh ectopic endometrial tissue and control endometrial tissue, and analyzed the expression of tRF&tiRNA in the two groups of exosomes through high-throughput sequencing

overall_design: Exosomes derived from ectopic endometrial tissue and control endometrial tissue

first: Yichen

last: Chen

first: Yingxue

last: Li


title: RNA-seq analysis of HIV-1 gp120 stimulated, p38α knockout and wild-type microglia isolated from mouse brain.

city: Riverside

first: Marcus

last: Kaul

address: 900 University Ave, SOM RB Room 311

department: Biomedical Sciences

institute: University of California, Riverside

summary: Purpose: The goals of this study is to compare transcriptome profiles of microglia from different mouse genotypes using RNA-seq analysis. HIV-associated neurocognitive disorders (HAND) occur in about 50% of infected individuals. Transgenic mice expressing the soluble HIV-1 envelope glycoprotein gp120 in astroglia (HIVgp120tg) display key neuropathological features of NeuroHIV patients (Toggas et al., 1994; Maung et al., 2014; Ojeda-Juarez et al., 2020). Here we show that microglial p38α MAPK plays a crucial role in neuronal injury triggered in vivo by the viral envelope glycoprotein. Cre-expression driven by the CX3CR1 promotor in HIVgp120tg mice carrying floxed alleles of p38a MAPK results in deletion of p38α in microglia and complete protection against neuronal injury and loss. RNA-seq analysis of microglia indicates that p38α deficiency leads to upregulation of neuroprotective and downregulation of neurotoxic factors. The expression patterns of genes associated with neurotransmission differ between brains protected from neuronal injury in the presence of transgenic gp120 and non-transgenic controls, suggesting a non-toxic modulation of neurons by the viral protein and microglial p38α deficiency.

overall_design: Examination of microglia gene expression in 4 different genotypes [wild type (WT), gp120-transgenic (gp120), microglia specific p38α knockout (p38α KO) and microglia specific p38α knockout gp120-transgenic (gp120 p38α KO)]. 3 Males and 3 Females samples were analyzed in each genotype.

first: Deepika

last: Bhullar

first: Ricky

last: Maung


title: Genome-wide maps of chromatin state and RNA-seq in WT and Dnmt3a KO BMDMs (bone marrow derived macrophages) [RRBS, ATAC-seq, CHIP-seq]

first: Peter

last: Kim

summary: To identify soluble factors released from Dnmt3a KO macrophages that might drive differences in osteoclast differentiation, we performed RNA sequencing on unstimulated BMDMs from WT and Dnmt3a KO mice. To investigate the mechanistic basis for the inflammatory phenotype. of myeloid cells with loss of Dnmt3a,we performed reduced representation bisulfite sequencing (RRBS) on Dnmt3a KO and WT BMDMs to assess methylation changes. Concurrently, we performed ATAC-sequencing (ATAC-seq) to assess differences in chromatin accessibility between WT and Dnmt3a KO BMDMs in the context of changes in methylation. We performed chromatin immunoprecipitation sequencing (CHIP-seq) for Irf3 and Rela based on ATAC-seq analysis.

overall_design: RRBS, ATAC-seq, CHIP-seq in WT and Dnmt3a KO BMDMs on day 8-10 of differentiation with or without LPS (10ng/ml for 6 hours for chromatin-related sequencing.

middle: G


title: PSAT1 drives extracellular vesicle release in multiple types of cancer and promotes bone metastasis by activating osteoclasts

city: Chuo-ku

first: Yusuke

last: Yamamoto

state: Tokyo

address: 5-1-1, Tsukiji, Chuo-ku

institute: National cancer center research institute

summary: Cancer cells secrete extracellular vesicles (EVs) to regulate cells in the tumor microenvironment for their own benefit to grow and survive in the patient’s body. Although emerging evidence has demonstrated the molecular mechanisms of EV release, regulating cancer-specific EV secretion remains challenging . In this study, we applied a microRNA (miRNA) library to reveal the universal mechanisms of EV secretion in cancer cells. Here, we identified miR-891b and its direct target gene, phospho-aminotransferase 1 (PSAT1), which promotes EV secretion through the serine-ceramide synthesis pathway. Inhibition of PSAT1 affected EV secretion in multiple types of cancer, suggesting that the miR-891b/PSAT1 axis shares a common mechanism of cancer EV secretion.

overall_design: Cancer cells (HCT116 and A549) were transfected with miR-891b or negative control by using Dharmafect1.

first: Tomofumi

first: Takahiro

last: Ochiya


title: Immediate Early Response 2 (Ier2) overexpression effect in NIH 3T3 cells

city: Mannheim

first: Carsten

last: Sticht

address: Theodor-Kutzer-Ufer

department: ZMF

institute: University Heidelberg

summary:
Analysis of Immediate Early Response 2 (Ier2)-inducible NIH 3T3 cells after Ier2 induction with RheoSwitch ligand RSL-1. Results provide insight into the function of Ier2 in NIH 3T3 mouse embryonal fibroblasts.
Immediate early genes, including Ier2, are rapidly induced in quiescent cells by proliferation and migration-inducing stimuli. Microarray gene expression profiling was performed to identify differentially expressed genes following overexpression of Ier2 in NIH 3T3-Ier2 inducible cells after 24 hour induction of Ier2.

overall_design: To prepare inducible NIH 3T3-RheoSwitch-Ier2 cells, NIH 3T3 cells were transfected with pNEBR-Ier2-hygro and pNEBR-R1 plasmids (Wallbaum et al., 2009, Biotechniques, 46: 433-440; PMID: 19480637). To induce Ier2 expression, NIH 3T3-RheoSwitch-Ier2 cells were treated with RheoSwitch ligand RSL-1.

first: Jonathan

last: Sleeman

first: Sabine

last: Wallbaum


title: Krüppel-Like Factor 5 Regulates Airway Epithelial Cell Identity and Function.

first: Ann

last: Harris

overall_design: Refer to individual Series


title: Activity-dependent modulation of synapse-regulating genes in astrocytes

city: La Jolla

first: Nicola

last: Allen

state: CA

address: 10010 North Torrey Pines Rd

institute: Salk Institute

summary: Analysis of astrocyte heterogeneity in the developing mouse visual cortex, and identification of genes regulated by neuronal and astrocyte activity

overall_design: Profile of glial cell nuclear mRNA at single cell level in P14 mouse visual cortex, from WT, VGlut2 cKO and Ip3r2 KO mice. 2 replicates for each genotype, each contains pooled tissue from 2 mice (4 biological replicates for each genotype total)

first: Isabella

last: Farhy-Tselnicker

first: Hanqing

last: Liu

first: Maxim

last: Shokhirev

first: Joseph

last: Ecker

middle: J


title: RNA sequencing of keratinocyte gene expression in keloid and normal skin tissue

status: Public on Oct 06 2021

city: Perth

first: Zhenjun

last: Deng

address: 35 Stirling Highway

institute: The University of Western Australia

summary: RNA sequencing has been performed to investigate the transcriptomic profile of keratinocyte derived from keloid and normal skin tissue. Keratinocytes of low passage (p2-4) were cultured onto T25 flasks until full confluence was reached. Keratinocytes were then harvested by trypsinization using 0.05% trypsin-EDTA and washed in Phosphate Buffered Saline (PBS). RNA was extracted using the RNeasy Mini kit protocol. Samples were then sent to the Australian Genome Research Facility (AGRF) where 1μg of RNA was submitted for next-generation sequencing. Once raw data returned, bioinformatics analyses were conducted. Through RNA-seq and bioinformatics analysis, 252 differentially expressed genes were identified in keloid keratinocytes compared to normal skin keratinocytes. Further gene ontology (GO) and Kyoto Encyclopaedia of Genes and Genomes (KEGG) analyses revealed that changes in tight junctions and dysregulated response to viral infection were associated with keloid epithelia and may be linked to the disease. The results of the transcriptome analysis intimate that keloid derived keratinocytes exhibit an abnormal gene expression profile and the abnormalities in keloid keratinocytes may have an important role in keloid pathogenesis.

overall_design: total RNA profiles of keloid keratinocytes and normal skin keratinocytes


title: UVB drives transcriptomic and epigenetic reprogramming which can be protected by antioxidant sulforaphane in human skin keratinocytes [RNA-seq]

first: Hsiao-Chen Dina

last: Kuo

overall_design: Samples of HaCaT cells were used for extraction of DNA and RNA; DNA was enriched by Agilent SureSelect Kit then bisulfite converted methyl-seq was performed. RNA-seq was performed with routine. Strategy. This dataset contains data for the RNA-seq only; Check the other SubSeries under this SuperSeries for Methyl-seq data.

first: Hsiao-Chen

first: Shanyi

last: Li

first: Lujing

last: Wang

first: Renyi

last: Wu

first: Davit

last: Sargsyan

first: Ah-Ng

last: Kong


title: A Prenylated dsRNA Sensor Protects Against Severe COVID-19

city: Glasgow

first: Marko

last: Noerenberg

address: Sir Michael Stoker Building

department: MRC-University of Glasgow Centre for Virus Research

institute: University of Glasgow

summary: iCLIP data to identify OAS1 binding sites in uninfected and SARS-CoV-2 infected cells

overall_design: For iCLIP analysis AAT-OAS1p46-RNase L CRISPR Guide 5 cells were either infected with SARS-CoV-2-CVR-GLA1 at MOI 2 or mock treated with medium only. After 24 hours incubation, cells were UV crosslinked and harvested for iCLIP. iCLIP was performed using an antibody specific for OAS1 and matched IgG control in addition to a size matched input.

first: Jeffrey

last: Lee

first: Natasha

last: Palmalux

first: Arthur

last: Wickenhagen


title: Effect of Nitidine Chloride on Rheumatoid Synovial Fibroblast-like Synoviocytes

first: Yu

last: Kuang

state: Guangdong

address: 58 Zhongshan 2nd Road, Yuexiu District

department: Rheumatology and Immunology

institute: The First Affiliated Hospital, Sun Yat-sen University

summary: To explore how nitidine chloride (NC) regulates biological functions of fibroblast-like synovioctyes (FLSs) from rheumatoid arthritis (RA) patients, we evaluated the transcriptome of NC-treated RA FLSs compared with untreated control using RNA sequencing analysis.

overall_design: Analysis of transcriptional changes in human RA FLSs treated with or without NC for 24h.

first: Chuyu

last: Shen

first: Shudi

last: Xu


title: Krüppel-Like Factor 5 Regulates Airway Epithelial Cell Identity and Function [CHIP-seq]

first: Ann

last: Harris

overall_design:
Examination of the cistrome of KLF5 and CEBPB in the Calu-3 cell line and KLF5 in the 16HBE14o- cell line

Please note that each processed data file was generated from both replicates and is linked to the corresponding rep1 sample records.

first: Alekh

last: Paranjapye

first: NandyMazumdar

last: Monali

first: Browne

last: James

first: Shih-Hsing

last: Leir


title: High-Throughput Single cell RNA sequencing of  Mesophyll cells from Maize

city: Nanjing

first: tao

middle: shen

last: tong

address: Weigang street 1st

institute: Nanjing Agricultural University

summary: We report the use of high-throughput single-cell RNA sequencing (scRNA-seq) to analyze gene expression in Mesophyll cells from Maize.  We using the 10x genomics platform to generate several millions of RNA-seq reads and enable transcriptional profiling of all genes of maize in a single-cell resolution. Tsne methods was used to perform dimension reduction analysis to visualize cells in a two-dimension axis.

overall_design: Single cell RNA sequencing for 14,656 cells (total) from maize (B73)

first: Wenli

last: Zhang

first: Lifen

last: Chen

first: Shentong

last: Tao


title: T-bet Suppresses TGF-b-independent Gut-resident Memory CD8+ T Cell Differentiation [ATAC-seq]

city: San Antonio

first: Nu

address: 7703 Floyd Curl Drive

institute: University of Texas Health Science Center at San Antonio

summary: TGF-beta signaling is required for the differentiation of gut-resident memory CD8 T cells. Here, we showed that the deficiency of transcription factor T-bet partially rescued the differentiation of TGF-beta receptor deficient gut-resident memory CD8 T cells.

overall_design:
Congenically marked naïve P14 TCR transgenic CD8 T cells were adoptively transferred into sex-matched C57BL/6 recipients followed by LCMV Armstrong infection. P14 T cells with different phenotypes were FACS sorted from both spleen and small intestine intraepithelial lymphocyte compartment.
Please note that processed data was generated from all replicates and is linked to the corresponding rep1 sample records.

first: Shruti

last: Mishra

first: Bingfei

last: Yu

first: Sara

last: Quon

first: Liwen

last: Wang

first: Chaoyu

last: Ma

first: Ananda

last: Goldrath


title: RNA-seq of in vitro differeentiated Th subsets and IL-27+TGF-β-treated CD4+ T cells

city: Bethesda

first: Hong-wei

last: Sun

address: 9000 Rockville Pile

department: Office of Science & Technology

institute: NIAMS

summary: IL-27 is an immunoregulatory cytokine whose essential function is to limit immune responses. We found that the gene encoding cholesterol 25-hydroxylase (Ch25h) was induced in CD4+ T cells by IL-27, enhanced by TGF-β, and antagonized by T-bet. Ch25h catalyzes cholesterol to generate 25-hydroxycholesterol (25OHC), which was subsequently released to the cellular milieu, functioning as a modulator of T cell response. Extracellular 25OHC suppressed cholesterol biosynthesis in T cells, inhibited cell growth, and induced nutrient-deprivation cell death without releasing high-mobility group box-1. This growth inhibitory effect was specific to actively proliferating cells with high cholesterol demand and was reversed when extracellular cholesterol was replenished. Interestingly, Ch25h expressing CD4+ T cells that received IL-27 and TGF-β signals became refractory to 25OHC-mediated growth inhibition in vitro. Nonetheless, IL-27 treated T cells negatively affected viability of bystander cells in a paracrine manner, but only if the bystander cells were in the early phases of activation. In mouse models of skin inflammation due to autoreactive T cells or chemically induced hypersensitivity, genetic deletion of Ch25h or Il27ra led to worsened outcomes. Thus, Ch25h is an immunoregulatory metabolic switch induced by IL-27 and dampens excess bystander T effector expansion in tissues through its metabolite derivative, 25OHC. This study reveals regulation of cholesterol metabolism as a modality for controlling tissue inflammation and thus represents a mechanism underlying T cell immunoregulatory functions.

overall_design: RNA-seq anslysis for in vitro differentiated CD4+ T cells. Naive CD4+ T cells were treated under the conditions (as descibed in the sample characteristics field) for three days, collected, and subjected to RNA-seq.

first: Hayato

last: Takahashi

first: Yohei

last: Mikami

first: Yuka

last: Kanno


title: Transcript profiling of human PBMC derived macrophage with or without Phago-Act treatment

city: Atlanta

first: Lei

last: Shi

state: GA

address: 100 Piedmont Avenue SE

department: Department of Biology

institute: Center for Diagnostics & Therapeutics

summary: To compare the gene expression of human macrophage with or without Phago-Act treatment, we performed a complete transcript profiling using transcriptome sequencing as a discovery platform.

overall_design: In each group, macrophages from 3 donors were used. Human PBMC derived macrophage were differenciated under MCSF for 5d, followed by Phago-Act treatment for 2d. Total RNAs were isolated from 3 samples (# 1-3) of human PBMC-derived SIRP low macrophage and donor-matched SIRPα normal macrophage.

first: Yuan

last: Liu


title: A genetic screen identifies a protective type III interferon response to Cryptosporidium that requires TLR3 dependent recognition

city: Philadelphia

first: Boris

last: Striepen

state: Pennsylvania

address: 380 S University Ave, Room H344

department: Pathobiology

institute: University of Pennsylvania

summary: Cryptosporidium is a leading cause of severe diarrhea and diarrheal-related death in children worldwide. As an obligate intracellular parasite, Cryptosporidium relies on intestinal epithelial cells to provide a niche for its growth and survival, but little is known about the contributions that the infected cell makes to this relationship. Here we conducted a genome wide CRISPR/Cas9 knockout screen to discover host genes required for Cryptosporidium parvum infection and/or host cell survival. The gene enrichment analysis indicated that the host interferon response, glycosaminoglycan (GAG) and glycosylphosphatidylinositol (GPI) anchor biosynthesis are important determinants of susceptibility to C. parvum infection. Several of these pathways are linked to parasite attachment and invasion and C-type lectins on the surface of the parasite. Evaluation of transcript and protein induction of innate interferons revealed a pronounced type III interferon response to Cryptosporidium in human cells as well as in mice. Treatment of mice with IFNλ reduced infection burden and protected immunocompromised mice from severe outcomes including death, with effects that STAT1 signaling in the enterocyte. Initiation of this type III interferon response was dependent on sustained intracellular growth and mediated by the pattern recognition receptor TLR3. We conclude that host cell intrinsic recognition of Cryptosporidium results in IFNλ production critical to early protection against this infection.

overall_design: This work includes two indepedent RNAseq experiments. The first is a 48 hour infection with C. parvum using either wild type parasites or transgenics propagated by our lab. The 48 hour experiment includes 3 biological replicates of uninfected, wild type infected, and transgenic infected. The second experiment was a 10 hour infection with a C. parvum strain expressing an mNeon green fluorescent protein. In this experiment cells were sorted to assess infection status with GFP negative as bystanders and GFP positive as infected cells. Three biological replicates were double sorted for these data.

first: Alexis

last: Gibson


title: UVB drives transcriptomic and epigenetic reprogramming which can be protected by antioxidant sulforaphane in human skin keratinocytes

city: Piscataway

first: Hsiao-Chen Dina

last: Kuo

address: 160 frelinghuysen road

institute: Rutgers University

summary: This SuperSeries is composed of the SubSeries listed below.


title: T-bet Suppresses TGF-b-independent Gut-resident Memory CD8+ T Cell Differentiation.

last: Zhang


title: Analysis of N6-methyladenosine Methylation Modification in Fructose-induced Non-alcoholic Fatty Liver Disease [RNA-seq2]

city: Shanghai

first: Yunchen

last: Luo

address: shanghai hongkou

institute: Shanghai Jiao Tong University School of Medicine

summary: Improvements in living standards have led to non-alcoholic fatty liver disease (NAFLD), one of the most common chronic liver diseases worldwide. Recent studies have shown that N6-methyladenosine (m6A), a type of RNA modification, is strongly associated with many important biological processes. However, the relationship between m6A methylation modifications and NAFLD remains poorly understood. In the present study, through methylated RNA immunoprecipitation sequencing and RNA transcriptome sequencing in high fructose diet-induced NAFLD mice, we found that hypermethylation-encoding genes were mainly enriched in lipid metabolism processes. We identified 266 overlapping and differentially expressed genes (DEGs) that changed at both the mRNA expression level and m6A modification level. Among them, 193 genes displayed increased expression and m6A modification, indicating that m6A RNA modifications tend to be positively correlated with NAFLD. We further compared the high fructose diet-induced NAFLD mouse model with leptin receptor-deficient mice and found that DEGs enriched in the lipid metabolism pathway were up-regulated in both groups. In contrast, DEGs associated with the immune inflammatory response were up-regulated in the high fructose diet group, but down-regulated in leptin receptor-deficient mice. Taken together, our results demonstrate that m6A methylation modifications may play an important role in the development of NAFLD.

overall_design: Male C57BL/6J mice aged 10-11 weeks were purchased from the Shanghai Laboratory Animal Company (SLAC, Shanghai, China). HFrD containing 10% kcal from fat, 70% kcal from carbohydrate, and 20% kcal from protein were purchased from Research Diets (D08040107, Research Diets, USA). After a 2-week intervention period in which mice were fed either a normal diet (ND) or HFrD, mice were anesthetized with 10% chloral hydrate.

first: Yan

last: Lu

first: Yongde

last: Peng